雌激素受体基因多态性与子宫内膜异位症关联研究

The study of the association of estrogen receptor gene polymorphisms with endometriosis

目的: 通过对雌激素受体基因多态性与子宫内膜异位症 (EM) 关系的研究, 明确遗传因素对EM的影响, 从分子遗传学的角度探讨EM的发病机制、早期诊断指标及方法。方法: 取 49名病理确诊为子宫内膜异位症的患者为EM组, 50名月经正常, 生育过正常小孩, 查无乳腺癌及妇科肿瘤, 在该院腹腔镜下或开腹做输卵管结扎或输卵管吻合术等, 排除EM、腺肌症的女性作为正常对照组。每位妇女取静脉血 2ml, 用酚 氯仿法提取和纯化DNA, 用设计雌激素受体 (ER) 基因类扩增ER基因 1号内含子及部分 2号外显子之间一段靶DNA引物, PCR变性、退火和延伸条件分别为 94℃30s、60℃40s、72℃90s,共 35个循环。PCR扩增产物用 1 5%agarose电泳检查扩增结果, 扩增产物大小为 1 3kb。PCR扩增后分别取 15UPvuⅡ与 10UXbaI内切酶于 37℃酶切至少 3h, 酶切产物经 1 5%琼脂糖溴乙锭电泳分离, 紫外灯下判断结果。结果: 标本经酶切后可产生 3种酶切格局: 使用PvuⅡ酶切可区分出 3种基因型: PP型 (终产物为一 1 3kb大小的条带), Pp型 (终产物为 1 .3kb、850bp、450bp大小的 3条带), pp型 (终产物为 850bp、450bp大小的 2条带), 使用XbaⅠ进行酶切可以区分出XX型 (1 .3kb), Xx型 (1. 3kb、910bp、390bp), xx型 (910bp、390bp) 3种基因型。

Objective:To evaluate whether the polymorphism in the ER gene is a potentially predisposing marker for endometriosis.This has important implications in terms of diagnosis and management of endometriosis at an early stage.Methods:About 49 patients who were diagnosed to have endometriosis were taken as study group.50 women who underwent health examination without endometriosis and gynaecological tumor were taken as control group for this study. Whole blood was collected and used for DNA isolation.DNA samples was amplified by PCR through 35 cycles. A cycling procedure included denaturation 30 seconds at 94 ℃, annealing for 40 seconds at 60 ℃ and primer extension for 90 secons at 72 ℃. After PCR amplification for ER gene, the PCR products were digested with restriction endonuclease, PvuⅡand XbaⅠ at 37 ℃ for at least three hours. Genotypes of the PvuⅡand XbaⅠ polymorphism were separated in 1.5% agarose gel electrophoresis and visualized by ethidium bromide staining. Results:PvuⅡ genotypes were defined as PP(1.3 kb), Pp(1.3 kb,850 bp,450 bp), or pp(850 bp,450 bp). XbaⅠgenotypes were defined as XX(1.3 kb), Xx(1.3 kb,910 bp,390bp), or xx(910 bp,390 bp).Conclusion:1.The polymorphism of the ER gene is not related to the development of endometriosis. 2.The frequencies of genotype Pp(0.44),pp(0.32),Xx(0.50),xx(0.42),ppxx (0.37) and PpXx(0.27) are much higher than others ,the genotype ppXX(0) is not found. the frequencies of the alleles p(0.54),x(0.67) of the ER gene are higher than P(0.46),X(0.33).